Novel formulations for the treatment of vaginal disorders

ABSTRACT

The present disclosure provides novel formulations suitable for the intravaginal delivery of tinidazole, as well as methods of using the same.

This application claims priority to U.S. Provisional Application No.61/839,070 filed Jun. 25, 2013, the entirety of which is incorporatedherein by reference.

NAMES OF THE PARTIES TO A JOINT RESEARCH AGREEMENT

For purposes of 35 U.S.C. 102(c), a joint research agreement wasexecuted between Mission Pharmacal and Université Laval in an inventionrelating to novel formulations for the treatment of vaginal disorders.

BACKGROUND OF THE INVENTION

Bacterial vaginosis (BV) is a common condition that is related toalterations in the normal vaginal flora. It is the most common cause ofvaginitis in women and has a recurrence rate of approximately 20%-40% atone month after therapy. In addition to the general discomfortassociated with the condition, BV has been linked to premature membranerupture, premature delivery, low birth weight, acquisition of HIV andother STDs, development of pelvic inflammatory disease (PID), andpost-operative infections following gynecological procedures.

Although the precise etiology of BV is unknown, the disease itself isassociated with the decrease or absence of protective lactobacilli whichare normally present in the vagina. Lactobacilli produce lactic acidfrom glycogen to maintain the vagina's acidic pH. This acidicenvironment inhibits the growth of other bacterial species typicallyfound in the vagina, albeit at low levels. Vaginal lactobacilli alsoproduce hydrogen peroxide (H₂O₂), which is toxic to viruses as well asto bacteria in vitro. When lactobacilli are sufficiently absent,bacteria such as Gardnerella vaginalis, Bacteroides spp., Mobiluncusspp., Haemophilus spp., peptostreptococci, Mycoplasma hominis,ureaplasma, and other anaerobes can populate the vaginal tract withoutdifficulty.

Currently approved treatments for BV include administering metronidazole(“MTZ”) 500 mg orally twice a day for 7 days; administering MTZ gel0.75%, one full applicator (5 g) intravaginally, once a day for 5 days;administering clindamycin cream 2%, one full applicator (5 g)intravaginally at bedtime for 7 days; administering tinidazole, 2 gorally once daily for 2 days; or administering tinidazole, 1 g orallyonce daily for 5 days.

Although MTZ remains the primary choice for treating BV in the UnitedStates, tinidazole is particularly useful for treating MTZ-resistant G.vaginalis, an organism commonly associated with BV. Despite thisutility, tinidazole must be dosed orally, whereas MTZ can be doseintravaginally. Common side effects associated with the oraladministration of tinidazole include, but are not limited to,metallic/bitter taste, nausea, anorexia, dyspepsia/cramps/epigastricdiscomfort, vomiting, constipation, tongue discoloration, stomatitis,diarrhea, decreased appetite, and flatulence.

Thus, there exists a need for novel tinidazole formulations suitable forthe treatment of BV and other diseases of the vaginal cavity thatrespond to or are susceptible to tinidazole (e.g. trichomoniasis) thatreduce or alleviate many of the side effects commonly associated withthe oral form of the drug. The present disclosure provides suchformulations.

BRIEF SUMMARY OF THE INVENTION

The present disclosure provides formulations suitable for theintravaginal delivery of tinidazole to a subject in need thereof, theformulations comprising water, tinidazole, a thermoreversible gellingagent, one or more pharmaceutically acceptable C₁-C₇ alcohols, asolubility enhancer, and, optionally, one or more preservatives.

In some embodiments, the thermoreversible gelling agent is triblockcopolymer having a central hydrophobic block flanked on each side with ahydrophilic block. In certain embodiments, the hydrophobic block ispolypropylene oxide). In some embodiments, the hydrophilic block ispoly(ethylene oxide).

In some embodiments, the thermoreversible gelling agent is poloxamer407.

In some embodiments, the one or more pharmaceutically acceptable C₁-C₇alcohols comprise a first pharmaceutically acceptable C₁-C₇ alcohol anda second pharmaceutically acceptable C₁-C₇ alcohol. In some embodiments,the first and second pharmaceutically acceptable C₁-C₇ alcohols areindependently selected from the group consisting of methanol, ethanol,isopropanol, propylene glycol, 2-(2-ethoxyethoxy)ethanol, benzylalcohol, and combinations thereof.

In some embodiments, the first pharmaceutically acceptable C₁-C₇ alcoholis benzyl alcohol.

In certain embodiments, the second pharmaceutically acceptable C₁-C₇alcohol is isopropanol.

In some embodiments, the isopropanol is a 60% (w/w) solution in water.

In some embodiments, the solubility enhancer is selected from the groupconsisting of mono- and di-alkyl ethers of isosorbide. In someembodiments, the mono- or di-alkyl ether of isosorbide is dimethylisosorbide.

In certain embodiments, the water comprises a mixture of steriledeionized water and a buffered aqueous solution. In certain embodiments,the buffered aqueous solution is citrate buffer.

In some embodiments, the formulation has a buffered pH selected from thegroup consisting of about 3 to about 5, about 3.5 to about 4.5, andabout 4 to about 4.5.

In certain embodiments, the tinidazole comprises from about 0.1 to about2% (w/w) of the formulations and in certain embodiments, the tinidazolecomprises about 1%, about 1.25%, or about 1.5% (w/w) of theformulations.

In some embodiments, the formulations comprise about 20% poloxamer 407(w/w), about 49.6% citrate buffer (w/w), about 3.1% benzyl alcohol(w/w), about 12% (w/w) of a 60% (w/w) solution of isopropanol in water,about 13.8% dimethylisosorbide (w/w), and about 1.5% tinidazole.

In other embodiments, the formulations comprise about 19% poloxamer 407(w/w), about 53% citrate buffer (w/w), about 2.9% benzyl alcohol (w/w),about 11% (w/w) of a 60% (w/w) solution of isopropanol in water, about12.7% dimethylisosorbide (w/w), and about 1.25% tinidazole.

In still further embodiments, the formulations comprise about 18%poloxamer 407 (w/w), about 57% citrate buffer (w/w), about 2.5% benzylalcohol (w/w), about 10% of a 60% (w/w) solution of isopropanol inwater, about 11.5% dimethylisosorbide (w/w), and about 1% tinidazole.

In other embodiments, the present disclosure provides methods fortreating bacterial vaginosis, trichomoniasis, or other diseases of thevaginal cavity susceptible to tinidazole, comprising intravaginallyadministering to a subject in need thereof a formulation comprisingwater, tinidazole, a thermoreversible gelling agent, one or morepharmaceutically acceptable C₁-C₇ alcohols, a solubility enhancer, and,optionally, one or more preservatives.

In certain embodiments of the methods described herein, thethermoreversible gelling agent is triblock copolymer having a centralhydrophobic block flanked on each side with a hydrophilic block. Incertain embodiments of the methods described herein, the hydrophobicblock is polypropylene oxide). In some embodiments of the methodsdescribed herein, the hydrophilic block is poly(ethylene oxide). Incertain embodiments of the methods described herein, thethermoreversible gelling agent is poloxamer 407.

In certain embodiments of the methods described herein, the one or morepharmaceutically acceptable C₁-C₇ alcohols comprises a firstpharmaceutically acceptable C₁-C₇ alcohol and a second pharmaceuticallyacceptable C₁-C₇ alcohol. In some embodiments of the methods describedherein, the first and second pharmaceutically acceptable C₁-C₇ alcoholsare selected from the group consisting of methanol, ethanol,isopropanol, propylene glycol, 2-(2-ethoxyethoxy)ethanol, benzylalcohol, and combinations thereof. In other embodiments of the methodsdescribed herein, the first pharmaceutically acceptable C₁-C₇ alcohol isbenzyl alcohol. In some embodiments of the methods described herein, thesecond pharmaceutically acceptable C₁-C₇ alcohol is isopropanol. Incertain embodiments of the methods described herein, the isopropanol isa 60% (w/w) solution in water.

In some embodiments of the methods described herein, the solubilityenhancer is selected from the group consisting of mono- and di-alkylethers of isosorbide. In some embodiments, the mono- or di-alkyl etherof isosorbide is dimethyl isosorbide.

In certain embodiments of the methods described herein, the watercomprises a mixture of sterile deionized water and a buffered aqueoussolution. In certain embodiments of the methods described herein, thebuffered aqueous solution is citrate buffer.

In some of the embodiments of the methods disclosed herein, theformulation has a buffered pH selected from the group consisting ofabout 3 to about 5, about 3.5 to about 4.5, and about 4.

In certain embodiments of the methods described herein, the tinidazolecomprises from about 0.1% to about 2% (w/w) of the formulation. In otherembodiments of the methods described herein, the tinidazole comprisesabout 1%, about 1.25%, or about 1.5% (w/w) of the formulation.

In certain embodiments of the methods described herein, the subject is ahuman.

The present disclosure also relates to a pharmaceutical compositioncomprising: about 18-20% poloxamer 407 (w/w), about 49.6-57% citratebuffer (w/w), about 2.5-3.1% benzyl alcohol (w/w), about 10-12% (w/w) ofa 60% (w/w) solution of isopropanol in water, about 11.5%-13.8%dimethylisosorbide (w/w), and about 1-1.5% tinidazole (w/w) (and/or anactive metabolite thereof), for treating bacterial vaginosis,trichomoniasis, or other disease(s) of the vaginal cavity susceptible totinidazole.

In certain embodiments the pharmaceutical composition exists as aviscous liquid at room temperature and gels at or close to bodytemperature.

The present disclosure further relates to the use of the formulations asdescribed herein for treating bacterial vaginosis, trichomoniasis, orother disease(s) of the vaginal cavity susceptible to tinidazole.

The present disclosure relates also to the use of the formulations asdescribed herein for the manufacture of a medicament for the treatmentof bacterial vaginosis, trichomoniasis, or other disease of the vaginalcavity susceptible to tinidazole.

An additional aspect of the present disclosure relates to a method fortreating a vaginal infection comprising intravaginally administering tothe subject a formulation containing about 0.1-2% tinidazole, or anactive metabolite thereof.

An additional aspect of the present disclosure relates to a method foradministering tinidazole to a human subject, comprising obtaining aformulation containing about 0.1-2% tinidazole, or an active metabolitethereof, wherein said formulation exists as a viscous liquid at roomtemperature; and intravaginally administering the formulation to a humansubject, wherein the formulation gels at or close to body temperature.

In a further embodiment, the present disclosure provides a formulationsuitable for the intravaginal delivery of tinidazole to a human subjectin need thereof, the formulation comprising tinidazole, wherein theformulation has a Franz cell flux of from about 2 to about 10(μmol/cm²)/√t(h).

In some embodiments, the Franz cell flux is from about 6 to about 9(μmol/cm²)/√t(h).

In another embodiment, the present disclosure provides a formulationsuitable for the intravaginal delivery of tinidazole to a human subjectin need thereof, the formulation comprising tinidazole, wherein theformulation has a mean flux rate in EpiVaginal™ tissue of from about 15nmol/cm²/min to about 30 nmol/cm²/min.

In certain embodiments, the mean flux rate in EpiVaginal™ tissue is fromabout 18 nmol/cm²/min to about 22.5 nmol/cm²/min.

The present disclosure also provides a formulation suitable for theintravaginal delivery of tinidazole to a human subject in need thereof,the formulations comprising tinidazole, wherein the formulation has amean tinidazole receiver fluid concentration at 1 hour of from about 0.5mM to about 2.5 mM.

In some embodiments, the formulation has a mean tinidazole receiverfluid concentration at 1 hour of from about 1.14 mM to about 1.74 mM.

The present disclosure also provides a tinidazole gel formulationsuitable for the intravaginal delivery of tinidazole to a subject inneed thereof, the formulation comprising water, tinidazole, athermoreversible gelling agent, one or more pharmaceutically acceptableC₁-C₇ alcohols, a solubility enhancer, and, optionally, one or morepreservatives, the tinidazole gel formulation prepared by the process ofa) dissolving the thermoreversible gelling agent in the water to form afirst solution; b) combining the one or more pharmaceutically acceptableC₁-C₇ alcohols, the solubility enhancer, and the tinidazole to form asecond solution; and c) adding the second solution to the firstsolution.

In certain embodiments, the thermoreversible gelling agent is triblockcopolymer having a central hydrophobic block flanked on each side with ahydrophilic block.

In certain embodiments, the hydrophobic block is polypropylene oxide).

In certain embodiments, the hydrophilic block is poly(ethylene oxide).

In certain embodiments, the thermoreversible gelling agent is poloxamer407.

In some embodiments, the one or more pharmaceutically acceptable C₁-C₇alcohols comprise a first pharmaceutically acceptable C₁-C₇ alcohol anda second pharmaceutically acceptable C₁-C₇ alcohol.

In some embodiments, the first and second pharmaceutically acceptableC₁-C₇ alcohols are independently selected from the group consisting ofmethanol, ethanol, isopropanol, propylene glycol,2-(2-ethoxyethoxy)ethanol, benzyl alcohol, and combinations thereof.

In certain embodiments, the first pharmaceutically acceptable C₁-C₇alcohol is benzyl alcohol.

In some embodiments, the second pharmaceutically acceptable C₁-C₇alcohol is isopropanol. In certain embodiments, the isopropanol is a 60%(w/w) solution in water.

In some embodiments, the solubility enhancer is selected from the groupconsisting of mono- and di-alkyl ethers of isosorbide. In certainembodiments, the mono- or di-alkyl ether of isosorbide is dimethylisosorbide.

In some embodiments, the water comprises a mixture of deionized waterand a buffered aqueous solution. In particular embodiments, the bufferedaqueous solution is citrate buffer.

In certain embodiments, the formulation has a pH of from about 3 toabout 5. In other embodiments, the formulation has a pH of from about3.5 to about 4.5.

The present disclosure further provides a process for the preparation ofa tinidazole gel formulation suitable for the intravaginal delivery oftinidazole to a human subject in need thereof, the process comprising a)dissolving a thermoreversible gelling agent in water to form a firstsolution; b) combining one or more pharmaceutically acceptable C₁-C₇alcohols, a solubility enhancer, and tinidazole to form a secondsolution; and c) adding the second solution to the first solution.

In certain embodiments, the thermoreversible gelling agent is triblockcopolymer having a central hydrophobic block flanked on each side with ahydrophilic block.

In some embodiments, the hydrophobic block is polypropylene oxide).

In some embodiments, the hydrophilic block is poly(ethylene oxide).

In some embodiments, the thermoreversible gelling agent is poloxamer407.

In some embodiments, the one or more pharmaceutically acceptable C₁-C₇alcohols comprise a first pharmaceutically acceptable C₁-C₇ alcohol anda second pharmaceutically acceptable C₁-C₇ alcohol.

In some embodiments, the first and second pharmaceutically acceptableC₁-C₇ alcohols are independently selected from the group consisting ofmethanol, ethanol, isopropanol, propylene glycol,2-(2-ethoxyethoxy)ethanol, benzyl alcohol, and combinations thereof.

In some embodiments, the first pharmaceutically acceptable C₁-C₇ alcoholis benzyl alcohol.

In certain embodiments, the second pharmaceutically acceptable C₁-C₇alcohol is isopropanol. In some embodiments, the isopropanol is a 60%(w/w) solution in water.

In certain embodiments, the solubility enhancer is selected from thegroup consisting of mono- and di-alkyl ethers of isosorbide. In someembodiments, the mono- or di-alkyl ether of isosorbide is dimethylisosorbide.

In some embodiments, the water comprises a mixture of deionized waterand a buffered aqueous solution. In certain embodiments, the bufferedaqueous solution is citrate buffer.

In some embodiments, the formulation has a mean tinidazole receiverfluid concentration at 2 hours of from about 1.5 mM to about 3.5 mM.

In other embodiments, the formulation has a mean tinidazole receiverfluid concentration at 4 hours of from about 3 mM to about 6 mM.

In other embodiments, the formulation has a mean tinidazole receiverfluid concentration at 2 hours of from about 2.3 mM to about 2.6 mM.

In some embodiments, the formulation has a mean tinidazole receiverfluid concentration at 4 hours of from about 4 mM to about 5.5 mM.

The present disclosure further provides a tinidazole gel formulationprepared by any of the processes described herein.

The present disclosure further provides a tinidazole gel formulationthat does not inhibit or only minimally inhibits lactobacillus strainsat a tinidazole concentration sufficient to inhibit Gardnerellavaginalis strains associated with bacterial vaginosis.

BRIEF DESCRIPTION OF THE DRAWINGS/FIGURES

The foregoing summary, as well as the following detailed description ofthe embodiments, will be better understood when read in conjunction withthe appended drawings. For the purpose of illustration, the drawings maydescribe the use of specific embodiments. It should be understood,however, that the formulations described herein are not limited to theprecise embodiments discussed or described in the figures.

FIG. 1 describes the standard for visually evaluating rabbit vaginaltissue exposed to the formulations described herein.

FIG. 2 describes the observed effects of vehicle formulation V1 onexternal rabbit vaginal tissue.

FIG. 3 describes the observed effects of vehicle formulation V2 onexternal rabbit vaginal tissue.

FIG. 4 describes the observed effects of vehicle formulation V3 onexternal rabbit vaginal tissue.

FIG. 5 describes the observed effects of formulation F1 on externalrabbit vaginal tissue.

FIG. 6 describes the observed effects of formulation F2 on externalrabbit vaginal tissue.

FIG. 7 describes the observed effects of formulation F3 on externalrabbit vaginal tissue.

FIG. 8 describes the evaluation standard for histopathological analysisof explanted rabbit vaginal tissue.

FIGS. 9A and 9B describe the observation and results ofhistopathological evaluation of rabbit vaginal explants previouslytreated with vehicle formulation V1.

FIGS. 10A and 10B describe the observation and results ofhistopathological evaluation of rabbit vaginal explants previouslytreated with vehicle formulation V2.

FIGS. 11A and 11B describe the observation and results ofhistopathological evaluation of rabbit vaginal explants previouslytreated with vehicle formulation V3.

FIGS. 12A and 12B describe the observation and results ofhistopathological evaluation of rabbit vaginal explants previouslytreated with formulation F1.

FIGS. 13A and 13B describe the observation and results ofhistopathological evaluation of rabbit vaginal explants previouslytreated with formulation F2.

FIGS. 14A and 14B describe the observation and results ofhistopathological evaluation of rabbit vaginal explants previouslytreated with formulation F3.

FIG. 15 is a graphical representation of the mean±SD of tinidazolereleased from formulations F1, F2, and F3 in Franz cell studies.

FIG. 16 is a graphical representation of a culture well and insert usedin the EpiVaginal™ tissue studies described herein.

FIG. 17 is an example of a multi-well culture plate representative ofthe types of multi-well culture plates that can be used in theEpiVaginal™ tissue studies described herein.

DETAILED DESCRIPTION OF THE INVENTION

The articles “a,” “an,” and “the” are used herein to refer to one or tomore than one (i.e., to at least one) of the grammatical object of thearticle. By way of example, “an element” means one element or more thanone element.

The phrase “substantially free of” as used herein means less than 5%,less than 2.5%, less than 1%, less than 0.1%, less than 0.01%, or lessthan 0.001% of a given component in the formulations described herein.

As used herein, “administration” or “administering” to a subjectincludes, but is not limited to, the act of a physician or other medicalprofessional prescribing a pharmaceutical composition of the inventionfor a subject.

Throughout the specification, the present disclosure provides certainranges and certain sub-ranges within those ranges. The identification ofcertain sub-ranges notwithstanding, the present disclosure should beread to include and disclose all sub-ranges within a given range (orsub-range). For example, if a range of 1 to 20 is described herein and asub-range of 5 to 15 is also described, the present disclosure should beunderstood to include and disclose all other sub-ranges within theoriginally defined range and sub-range, e.g., 1 to 5, 7 to 12, 15 to 20,2 to 18, etc.

The present disclosure provides novel aqueous tinidazole formulationssuitable for intravaginal administration to a subject in need thereof.In particular embodiments, the formulations can comprise water,tinidazole or an active metabolite thereof, a thermoreversible gellingagent, one or more pharmaceutically acceptable C₁-C₇ alcohols, asolubility enhancer, and, optionally, one or more preservatives. Incertain embodiments, the formulations can further include a polyol, suchas, but not limited to polyethylene glycol, an optional penetrationenhancer, or an optional excipient such as HPMC. In particularembodiments, the formulations can be completely or substantially free ofboric acid and/or ethylenediaminetetraacetic acid (EDTA). In certainembodiments, the formulations can be free or substantially free of asecond active ingredient, such that tinidazole is the only activeingredient in the formulations.

An important characteristic of the formulations described herein aretheir thermoreversible nature. In particular embodiments, theformulations exist as a viscous liquid at room temperature (i.e. about20° C. to about 23° C.). At higher temperatures, though, theformulations can exist as a gel. In particular embodiments, theformulations can be a gel at or close to body temperature (i.e. about35° C. to about 37° C.). Without wishing to be bound to any particulartheory, it is believed that gel formation at higher temperature is auseful property because it allows the formulations to flow into thevaginal cavity and to reach the smallest irregularities of the mucosalsurface where they subsequently gel and adhere to biological surfacessuch as the vaginal epithelia.

Although using thermoreversible gels for the prevention of STDs waspreviously disclosed, see, e.g. WO 97/42962 and WO 99/53897 (both ofwhich are hereby incorporated by reference in their entirety), neitherpublication teaches the use of tinidazole. Moreover it has now beenunexpectedly discovered that the percentage of tinidazole in aformulation including a thermoreversible gelling agent can besignificantly increased using the formulations disclosed herein.

In certain embodiments, tinidazole can be used in the formulationsdescribed herein at any suitable concentration. In other embodiments,the concentration of tinidazole can be about 0.1% to about 2% (w/w),about 0.5% to about 2% (w/w), or about 0.75% to about 2% (w/w). Inparticular embodiments, the concentration of tinidazole can be about0.75 to about 1.5% (w/w). In specific embodiments, the concentration oftinidazole can be about 1%, about 1.25%, or about 1.5% (w/w). Theformulations can also include an active metabolite of tinidazole at anysuitable concentration. Known active metabolites of tinidazole include,but are not limited to1-(2-(ethylsulfonyl)ethyl)-2-methyl-4-nitro-1H-imidazol-5-ol and2-hydroxymethyl tinidazole.

In certain embodiments, the thermoreversible gelling agent can bepresent in the formulations described herein at any suitableconcentration. In certain embodiments, the thermoreversible gellingagent can be at a concentration of about 5% to about 50% (w/w) and incertain embodiments, at a concentration of about 15% to about 35% (w/w).In particular embodiments, the thermoreversible gelling agent can bepresent at a concentration of about 15% to about 25% (w/w). In certainembodiments, the thermoreversible gelling agent can be present in anamount ranging from about 15% to about 25% (w/w); in an amount rangingfrom about 17% to about 22% (w/w); or in an amount ranging from about18% to about 20% (w/w). In certain embodiments, the thermoreversiblegelling agent can be present at a concentration of about 18% (w/w),about 19% (w/w), or about 20% (w/w). Exemplary thermoreversible gellingagents include, but are not limited to, triblock copolymers having acentral hydrophobic block flanked on each side with a hydrophilic block.In certain embodiments the hydrophobic block can be a block ofpolypropylene oxide). In certain embodiments, the hydrophilic blocks canbe blocks of poly(ethylene oxide). In particular embodiments, thethermoreversible gelling agent can be selected from the group consistingof poloxamer 407 (CAS 9003-11-6), poloxamer 124, poloxamer 188,poloxamer 237, poloxamer 338. In specific embodiments, thethermoreversible gelling agent is poloxamer 407.

The one or more pharmaceutically acceptable C₁-C₇ alcohols can bepresent in the formulations at any suitable concentration and caninclude any appropriate straight chain, branched, cyclic, or aromaticalcohol having the designated number of carbons. Exemplarypharmaceutically acceptable C₁-C₇ alcohols include, but are not limitedto, ethanol, isopropanol, propylene glycol, 2-(2-ethoxyethoxy)ethanol,and benzyl alcohol. In certain embodiments, the one or morepharmaceutically acceptable C₁-C₇ alcohols can be present at aconcentration of about 2% to about 20% (w/w). In other embodiments, theone or more pharmaceutically acceptable C₁-C₇ alcohols can be present ata concentration of about 5% to about 15% (w/w). In particularembodiments, the one or more pharmaceutically acceptable C₁-C₇ alcoholscan be present from about 8% to about 11% (w/w).

In certain embodiments, the one or more pharmaceutically acceptableC₁-C₇ alcohols can include one to ten, i.e. 1, 2, 3, 4, 5, 6, 7, 8, 9 or10 pharmaceutically acceptable C₁-C₇ alcohols. In certain embodiments,the formulations can include a first C₁-C₇ alcohol and a second C₁-C₇alcohol. In certain embodiments the first C₁-C₇ alcohol can be presentat a concentration of about 2.25% to about 3.25% (w/w). In certainembodiments, the first C₁-C₇ alcohol can be present in an amount ofabout 2.5%, about 2.9%, or about 3.2% (w/w). In other embodiments, thesecond C₁-C₇ alcohol can be present at a concentration of about 10% toabout 12% (w/w), and in certain embodiments can be about 6%, about 6.5%,or about 7.2% (w/w) of the formulation. In certain embodiments, thefirst C₁-C₇ alcohol can be benzyl alcohol. In certain embodiments, thesecond C₁-C₇ alcohol can be isopropyl alcohol. In certain embodiments,the isopropyl alcohol can be a 60% solution (w/w) in water.

The formulations can also include a solubility enhancer, useful forenhancing the solubility of tinidazole in the formulation. Exemplarysolubility enhancers include, but are not limited to, mono and di-alkylethers of isosorbide. In particular embodiments, the solubility enhanceris dimethylisosorbide (“DMI”). The solubility enhancer can be present inthe formulations at any suitable concentration; however, in certainembodiments it can be present at a concentration of about 10% to about15% (w/w). In other embodiments, the solubility enhancer can be presentat a concentration of about 11.5%, about 12.7%, or about 14% (w/w).

The formulations can further include an optional penetration enhancer.The optional penetration enhancer can be used in the formulations in anysuitable concentration; however in certain embodiments, it can bepresent in the formulations in a range of about 0.01% to 15% (w/w).Exemplary penetration enhancers include, but are not limited to terpenesand terpenoids (such as menthol and the like); pyrrolidones (such asN-methyl-2-pyrrolidone, 1-dodecylazacycloheptan-2-one, and the like);sulfoxides (such as DMSO and the like); phospholipids; cyclodextrinssuch as, but not limited to, O-cyclodextrin; dodecyl-N,N-dimethylaminoacetate; clofibric acid; and amino acid derivatives such as, but notlimited to, dodecyl N,N-dimethylamino isopropionate. While in certainembodiments the optional penetration enhancer can be present, in otherembodiments, the penetration enhancer can be completely or substantiallyabsent.

The formulations can optionally include one or more preservatives in apharmaceutically acceptable amount. Suitable exemplary preservativesinclude, but are not limited to, methylparaben, propylparaben, BHA, BHT,and combinations of the foregoing. In certain embodiments, theformulations can comprise less than about 1% (w/w), less than about 0.5%(w/w), or less than about 0.05% (w/w) of the formulation.

In typical embodiments, the remainder of the formulations compriseswater. As used herein “water” refers to sterile deionized water,buffered aqueous solutions comprising sterile deionized water, or anycombination of the foregoing. The amount of water in the formulationscan be at least about 10%, 20%, 30%, 40%, 45%, 47%, 50%, 55%, 58%, 60%,61%, 65% or at least about 70% (w/w). In certain embodiments, the amountof water in the formulations can range from about 40% (w/w) to about 70%(w/w), from about 50% to about 70% (w/w), or from about 50% to about 65%(w/w). In particular embodiments, the amount of water in theformulations can be about 54.4% (w/w), about 57.6% (w/w), or about 61%(w/w).

Buffered aqueous solutions can contain one or more pharmaceuticallyacceptable buffering agents suitable for maintaining the pH of theformulations in a range of about 3 to about 5, and in certainembodiments, about 3.5 to about 4.5. Suitable buffering agents andsystems are well known to those of ordinary skill in the art; however incertain embodiments, the buffering agent is a citrate buffer. Otherpossible buffering agents include, but are not limited to, veronalacetate buffer, and acetate buffer. In particular embodiments, the pH ofthe formulations is buffered to a pH of about 4 to about 4.5.

The viscosity of the formulations can range from about 1 cP to about100,000 cP, about 1 cP to about 50,000 cP, about 1 cP to about 10,000cP, or about 1 cP to about 1,000 cP in its ungelled state when measuredusing a Brookfield DV-II+Pro Extra LV™ viscometer with spindle type 63at 100 RPM at about 21° C. to about 22° C. In particular embodiments,the ungelled formulations can have a viscosity of about 1 to about 900cP. In certain embodiments, the ungelled formulations can have aviscosity of about 200 cP to about 750 cP, and in other embodiments, theungelled formulations can have a viscosity of about 300 cP to about 550cP. In particular embodiments, the composition can have a viscosity ofabout 315 cP to about 345 cP. In other embodiments, the ungelledformulations can have a viscosity of about 420 cP to about 460 cP. Instill other embodiments, the formulations can have a viscosity in theungelled state of about 480 cP to about 550 cP.

In certain embodiments, the formulations can have a temperature ofgelation (T_(gel)) of about 18 to about 37° C. In other embodiments, theformulations can have a T_(gel) of about 22 to about 35° C. In stillfurther embodiments, the formulations can have a T_(gel) of about 30° C.to about 35° C. And in still other embodiments, the formulations canhave a T_(gel) of about 32° C.

In addition to providing the formulations discussed above, the presentdisclosure also provides methods for treating BV, trichomoniasis, orother disease or disorder suitable for treatment via the intravaginaladministration of a tinidazole. Suitable methods for treating BV includeintravaginally administering the formulations to a subject in needthereof. Exemplary subjects include mammals, preferably humans. Incertain embodiments, the formulations can be administered intravaginallyonce a day for a period of up to about 30 days. However, in otherembodiments, the formulations can be administered chronically over aperiod of months or years.

In certain embodiments, the formulations can be administeredintravaginally twice a day for a period of up to about 30 days. Inspecific embodiments, the formulations can be administeredintravaginally once or twice a day for a period of 1, 2, 3, 4, 5, 6, 7,8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25,26, 27, 28, 29, or 30 days. In some embodiments, the formulations can beadministered intravaginally once or twice a day every other day, once ortwice a day every third day, once or twice a day every fourth day, onceor twice a day every fifth day, once or twice a day every sixth day,once or twice a day once a week, once or twice a day one time everyother week, once or twice a day one time every third week, once or twicea day once a month. In certain embodiments, the formulations can beadministered intravaginally in a single application, i.e., in thesubstantial absence of further dosing until such time as symptoms recuror until the subject is directed by a physician to resume treatment.

The present formulations also have surprising stability at lowtemperature. For example, in certain embodiments, the formulations willshow little or no tinidazole crystallization when chilled to as low asabout 6° C. for 24 hours and then warmed to 22 C for 24 hours. In otherembodiments, the formulations will show little or no tinidazolecrystallization when chilled to as low as about 14° C. for 24 hours andthen warmed to 22° C. for 24 hours.

The present formulations similarly exhibit unexpected long-termstability at 22° C., showing little or no tinidazole crystallization at1, 2, and in certain embodiments 2.6 or 2.8 months. In other embodimentsthe formulations can be stable for at least about 10 months, at leastabout 12 months, and in other embodiments, at least about 18 months.

The present formulations have similarly shown a surprising lack ofirritation in a rabbit vaginal model, as described in detail below.

In addition to the properties noted above, the formulations describedherein can exhibit a “Franz cell flux.” As used herein, a “Franz cellflux” refers to the rate at which tinidazole in the formulationsdescribed herein crosses a polycarbonate membrane having 0.40 μm poresand an area of 1.77 cm² in a Franz cell diffusion study at 37° C.wherein the receiver fluid is 0.05 M citrate buffer (pH 3.3). In someembodiments, Franz cell flux can range from about 2 to about 15(μmol/cm²)/√t, where t is measured in hours (“√t(h)”). In otherembodiments, the Franz cell flux can range from about 4 to about 10(μmol/cm²)/√t. In still further embodiments, the Franz cell flux canrange from about 6 to about 9 (μmol/cm²)/√t(h). In particularembodiments, the Franz cell flux can be about 6, about 7.5, or about 8.7(μmol/cm²)/√t(h). In particular embodiments, the formulations can be anyof formulations F1, F2, or F3 described elsewhere herein.

In other embodiments, Franz cell flux can range from 2 to 15(μmol/cm²)/√t(h). In other embodiments, the Franz cell flux can rangefrom 4 to 10 (μmol/cm²)/√t(h). In still further embodiments, the Franzcell flux can range from 6 to 9 (μmol/cm²)/√t(h). In particularembodiments, the Franz cell flux can be 6, 7.5, or 8.7 (μmol/cm²)/√t(h).In particular embodiments, the formulations can be any of formulationsF1, F2, or F3 described elsewhere herein.

The formulations described herein can further exhibit a mean flux ratein EpiVaginal™ tissue culture. The “mean flux rate in EpiVaginal™ tissueculture” refers to the rate at which tinidazole in a 100 μL aliquot ofthe formulations described herein at 37° C. crosses a full thicknesstissue culture produced from normal human-derived vaginal-ectocervicalepithelial cells (VEC-100-FT EpiVaginal™ tissue available from MatTekCorporation, Ashland, Mass.), mounted on polycarbonate membranes having0.4 μm pores, wherein the culture medium (alternatively referred to asthe receiver fluid) is 0.5 ml VEC-100-ASY available from MatTekCorporation (Ashland, Mass.).

In particular embodiments, the mean flux rate (nmol/cm²/min) can rangefrom about 15 nmol/cm²/min to about 30 nmol/cm²/min. In otherembodiments, the mean flux rate can range from about 15 nmol/cm²/min toabout 25 nmol/cm²/min. In other embodiments, the mean flux rate canrange from about 17 nmol/cm²/min to about 25 nmol/cm²/min. In otherembodiments, the mean flux rate can range from about 18 nmol/cm²/min toabout 22.5 nmol/cm²/min. In particular embodiments, the mean flux ratecan be about 18, about 19, or about 22.5 nmol/cm²/min.

In other embodiments, the mean flux rate (nmol/cm²/min) can range from15 nmol/cm²/min to 30 nmol/cm²/min; from 15 nmol/cm²/min to 25nmol/cm²/min; from 17 nmol/cm²/min to 25 nmol/cm²/min; or from 18nmol/cm²/min to 22.5 nmol/cm²/min. In particular embodiments, the meanflux rate can be 18, 19, or 22.5 nmol/cm²/min.

The formulations described herein can also have a mean tinidazolereceiver fluid concentration. The “mean tinidazole receiver fluidconcentration” is the concentration of tinidazole in receiver fluid at agiven time point after application of a 100 μL aliquot of theformulations described herein at 37° C. to full thickness tissue cultureproduced from normal human-derived vaginal-ectocervical epithelial cells(VEC-100-FT EpiVaginal™ tissue available from MatTek Corporation,Ashland, Mass.), mounted on polycarbonate membranes having 0.4 μm pores,wherein the culture medium/receiver fluid is VEC-100-ASY available fromMatTek Corporation (Ashland, Mass.).

In particular embodiments, the mean tinidazole receiver fluidconcentration after 1 hour can be about 0.5 mM to about 2.5 mM. In otherembodiments, the mean tinidazole receiver fluid concentration after 1hour can be about 1 mM to about 2 mM. In still other embodiments, themean tinidazole receiver fluid concentration after 1 hour can be about1.14 mM to about 1.74 mM. In particular embodiments, the mean tinidazolereceiver fluid concentration after 1 hour can be about 1.14 mM, about1.5 mM, or about 1.74 mM.

In other embodiments, the mean tinidazole receiver fluid concentrationafter 1 hour can range from 0.5 mM to 2.5 mM; from 1 mM to 2 mM; or from1.14 mM to 1.74 mM. In particular embodiments, the mean tinidazolereceiver fluid concentration after 1 hour can be 1.14 mM, 1.5 mM, or1.74 mM.

In particular embodiments, the mean tinidazole receiver fluidconcentration after 2 hours can be about 1.5 mM to about 3.5 mM. Inother embodiments, the mean tinidazole receiver fluid concentrationafter 2 hours can be about 2 mM to about 3 mM. In still otherembodiments, the mean tinidazole receiver fluid concentration after 2hours can be about 2.3 mM to about 2.6 mM. In particular embodiments,the mean tinidazole receiver fluid concentration after 2 hours can beabout 2.3 mM, about 2.9 mM, or about 2.6 mM.

In other embodiments, the mean tinidazole receiver fluid concentrationafter 2 hours can range from 1.5 mM to 3.5 mM; from 2 mM to 3 mM; orfrom 2.3 mM to 2.6 mM. In particular embodiments, the mean tinidazolereceiver fluid concentration after 2 hours can be 2.3 mM, 2.9 mM, or 2.6mM.

In other embodiments, the mean tinidazole receiver fluid concentrationafter 4 hours can be about 3.5 mM to about 6 mM. In other embodiments,the mean tinidazole receiver fluid concentration after 4 hours can beabout 4 mM to about 5.5 mM. In particular embodiments, the meantinidazole receiver fluid concentration after 4 hours can be about 4.4mM, about 5.6 mM, or about 5.1 mM.

In other embodiments, the mean tinidazole receiver fluid concentrationafter 4 hours can range from 3.5 mM to 6 mM or from 4 mM to 5.5 mM. Inparticular embodiments, the mean tinidazole receiver fluid concentrationafter 4 hours can be 4.4 mM, 5.6 mM, or 5.1 mM.

The formulations described herein can also have a concentrationdependent mean peak tinidazole concentration in EpiVaginal™ tissue(“mean peak concentration”). The mean peak concentration is the averagemaximum concentration of tinidazole in full thickness tissue cultureproduced from normal human-derived vaginal-ectocervical epithelial cells(VEC-100-FT EpiVaginal™ tissue available from MatTek Corporation,Ashland, Mass.) after application of a 100 μL aliquot of theformulations described herein at 37° C. for a period of time sufficientto achieve the maximum tinidazole concentration in the tissue, whereinthe tissue is mounted on polycarbonate membranes having 0.4 μm pores,wherein the culture medium/receiver fluid is VEC-100-ASY available fromMatTek Corporation (Ashland, Mass.).

In particular embodiments, the mean peak concentration can range fromabout 1500 ng tinidazole/mg tissue to about 4500 ng tinidazole/mgtissue. In other embodiments, the mean peak concentration can range fromabout 1800 to about 4200 ng tinidazole/mg tissue. In still furtherembodiments, the mean peak concentration can range from about 2000 ngtinidazole/mg tissue to about 4000 ng tinidazole/mg tissue. In certainembodiments, the mean peak concentration can be about 2000 ngtinidazole/mg tissue, about 2700 ng tinidazole/mg tissue, or about 3400ng tinidazole/mg tissue.

In other embodiments, the mean peak concentration can range from 1500 ngtinidazole/mg tissue to 4500 ng tinidazole/mg tissue. In otherembodiments, the mean peak concentration can range from 1800 to 4200 ngtinidazole/mg tissue. In still further embodiments, the mean peakconcentration can range from 2000 ng tinidazole/mg tissue to 4000 ngtinidazole/mg tissue. In certain embodiments, the mean peakconcentration can be 2000 ng tinidazole/mg tissue, 2700 ng tinidazole/mgtissue, or 3400 ng tinidazole/mg tissue.

In a further embodiment, the formulation can have a mean peak tinidazoleconcentration that satisfies the formula:

y=2834x−804.83

over a range of tinidazole concentrations of from about 0.8% (w/w) toabout 1.8% (w/w), wherein y is the mean peak concentration of tinidazolemeasured in the culture and x is the concentration of tinidazole in theformulation.

EXAMPLES

The formulations and methods are now further detailed with reference tothe following examples. These examples are provided for the purpose ofillustration only and the formulations and methods described hereinshould in no way be construed as being limited to these examples.Rather, the formulations should be construed to encompass any and allvariations which become evident as a result of the teaching providedherein.

The 60% IPA solution (w/w) described below was prepared by adding 600 gof IPA USP and 400 g of water USP to a 1.0 L vessel and mixing thecombined solution well.

The 0.05 M citric acid buffer solution used and described in theexamples below was prepared as follows. 21.016 g of citric acidmonohydrate was added to a 2.0 L vessel with 2000 mL of water (“solution1”). In a separate 2 L vessel, 25.806 g of sodium citrate (anhydrous)was mixed with 2000 mL of water until the material was dissolved(“solution 2”). Finally, 550 mL of solution 1 and 450 mL of solution 2were combined and mixed in an appropriately sized vessel. The pH of theresulting solution was measured at 3.30 using a standard pH electrode.

Example 1 Timidazole Gel Formulations

Formulations having 1%, 1.25%, and 1.5% tinidazole as shown in Table 1,below, were prepared according to the following general procedure.

An appropriate quantity of poloxamer 407 and citrate buffer werecombined in an appropriate vessel to give a suspension. The vessel wasthen placed in an ice-water bath and the suspension was mixed at 400 rpmfor 6 hours to dissolve all of the poloxamer. Upon complete dissolution,the clear solution was warmed to room temperature (22° C.) and thistemperature was maintained with a water bath.

Next, the appropriate quantities of 60% IPA, benzyl alcohol, DMI andtinidazole were combined in an appropriately sized vessel. The resultingmixture was then heated at 37° C. for approximately 30 minutes until allof the tinidazole was dissolved. Once dissolved, the solution was thencooled to room temperature and added slowly and with agitation to thepoloxamer solution. The resulting mixture was then stirred at low RPMuntil uniform. After stirring, the mixing apparatus was removed and theresulting material was allowed to settle for 24 hours to allow trappedair to bubble to the surface.

TABLE 1 Citrate Benzyl Formu- Poloxamer Buffer Alcohol 60% IPA DMITinidazole lation 407 (g)/(% (g)/(% (g)/(% (g)/(% (g)/ (g)/(% # w/w)w/w) w/w) w/w) (% w/w) w/w) F1 10.00 24.79 1.56 6.00 6.90 0.750 (20.00)(49.58) (3.12) (12.00) (13.80) (1.50) F2 9.50 26.62 1.43 5.50 6.33 0.625(19.00) (53.24) (2.86) (11.00) (12.6) (1.25) F3 9.00 28.50 1.25 5.005.75 0.500 (18.00) (57.00) (2.50) (10.00) (11.50) (1.00)

Example 2

Formulations 4 and 5, described in Table 2, below, were preparedaccording to the following general procedure.

An appropriate quantity of poloxamer 407 and citrate buffer werecombined in an appropriate vessel to give a suspension. The vessel wasthen sealed and placed on a shaker rotating at ˜160 rpm in a cold roomat 4° C. and was left shaking overnight until complete dissolution ofthe poloxamer was observed. The resulting clear solution was allowed tostand at 4° C. for a few hours to remove air bubbles present in thesolution. The poloxamer solution was then placed in an incubator at 18°C. for approximately 30 min.

Next, the appropriate quantities of DMI, 70% IPA, benzyl alcohol, andtinidazole were combined in an appropriately sized vessel. The resultingmixture was heated at 37° C. for approximately 10 min until completedissolution of the tinidazole was observed. The clear solution was thencooled to room temperature (22° C.) and was added slowly with agitation(shaking) to the poloxamer solution at room temperature (22° C.) until auniform clear solution was obtained. The final formulations were allowedto stand at room temperature (22° C.) to remove air bubbles.

TABLE 2 26% Poloxamer Benzyl Citrate Formulation Gel Soln DMI 70% IPAAlcohol Buffer Tinidazole Tinidazole # (g) (ml) (ml) (ml) (ml) (mg) % F414.23 2.2 2.4 0.55 0.37 250 1.25 F5 14.23 2.0 2.2 0.50 0.87 200 1.00

The cold temperature stability of formulations 4 and 5 was tested bycooling the formulations to a specified temperature for 24 hours andthen warming the formulations to room temperature (22° C.) where theywere allowed to stand for a further 24 hours. During the temperaturechallenge, the formulations were visually evaluated for the formation ofcrystals either at low temperature or upon subsequent warming. Theresults are shown in Table 3.

TABLE 3 Temp (° C.) Formulation # 18 14 10 6 F4 No crystals No crystalsCrystals Crystals F5 No crystals No crystals No crystals No crystals

This data demonstrates that the formulations can be subjected torelatively cool temperatures and maintain tinidazole in solution, evenat high tinidazole concentrations.

Example 3 In Vitro Drug Release

To determine whether tinidazole would be released upon intravaginaladministration, an in vitro test (membrane-free model) simulatingformulation erosion in the vaginal cavity was conducted. Two differenttinidazole 1.25% gel formulations were prepared (F6=18.6% gel, 1.75%hydroxypropyl methylcellulose (HPMC), Tmelt=31° C.; F7=19.01% gel, noHPMC, T_(m)=29° C.). Specifically, 1 g of each formulation was added toa pre-weighed vial equilibrated at 37° C. Next, 0.5 ml of deionizedwater was slowly added to the top of the vial and the vial was allowedto stand for 2 hours, 6 hours, 24 hours, or 4 days. After theappropriate amount of time passed, the liquid supernatant sitting on topof the formulation was carefully removed, weighed, and analyzed for itstinidazole concentration using standard HPLC methodology. All studieswere conducted in triplicate to ensure quality data. This membrane-freesystem is a suitable choice to mimic natural gel erosion in the vaginalcavity.

These studies showed that the formulations described herein aresusceptible to gradual erosion/dissolution with minimal or no agitation.

Example 4 Vaginal Irritation in Rabbits

Formulations F1, F2, and F3 (1 mL) (as described in Example 1) andVehicle Formulations V1, V2, and V3 (as described below in Table 4) wereeach administered into the vaginal vaults of 3 female test rabbits (NewZealand White strain) for five consecutive days. The formulations wereadministered with a blunt tipped animal-feeding needle attached to a 3cc syringe. Control animals were dosed with 1 mL of a 0.9% salinesolution under identical conditions.

TABLE 4 Vehicle Formulations Citrate Benzyl Poloxamer Buffer Alcohol 60%Tinidazole (g)/ (g)/(% (g)/(% IPA (g)/ DMI (g)/ (g)/(% (% w/w) w/w) w/w)(% w/w) (% w/w) w/w) V1 10.00 40.00 N/A N/A N/A N/A (20.00) (80.00) V210.00 25.54 1.56 6.00 6.90 N/A (20.00) (51.10) (3.12) (12.00) (13.80) V38.75 30.45 1.13 4.50 5.18 N/A (17.50) (60.90) (2.26) (9.00) (10.36)

Prior to the first dose of the control, vehicle, or active formulations,and before each daily dose thereafter, the external vaginal tissues ofeach animal was observed for erythema, exudate, and edema according tothe scoring system shown in FIG. 1. The results of the daily monitoringof the external vaginal tissue are shown in FIGS. 2 (V1), 3 (V2), 4(V3), 5 (Formulation 1), 6 (Formulation 2), and 7 (Formulation 3). Ascan be seen in the Figures, each of the formulations caused minimal orno visual changes in the external vaginal tissue of the subject animals.

Following completion of the protocol, the animals were sacrificed withsodium pentobarbital and the vaginal tissue of each animal was carefullyremoved. The tissue was then observed macroscopically and scored forirritation and injury to the epithelial layer. Following macroscopicobservation, the tissue was sectioned to provide the cervical, central,and caudal portions. The samples were then individually placed in 10%neutral buffered formalin and submitted for histopathologicalevaluation. Samples were evaluated based on the criteria shown in FIG.8.

Specific results, corresponding to formulations V1, V2, V3, F1, F2, andF3 are shown in FIGS. 9A and 9B through FIGS. 14A and 14B, respectively.Overall, each of the vehicle and drug-containing formulations showedminimal observable effects, suggesting that the formulations aresuitable for intravaginal administration despite the propensity for theindividual components of the formulations to cause irritation.

Example 5 Isolation and Identification of Bacterial Strains Associatedwith Bacterial Vaginosis

To obtain clinical isolates from women suspected of having bacterialvaginosis, 192 microscope slides from 192 vaginal swabs (obtained fromwomen consulting in the Quebec City area) submitted for Nugent scoringwere analyzed. Based on microscopic observations of bacterial cellmorphology and Nugent score, 46 swabs were selected for bacterialisolation.

Bacterial isolates were identified by polymerase chain reaction (PCR)and DNA sequencing of the 16S rRNA gene. A total of 30 swabs werepositive for at least one isolate of Bacteroides spp., Gardnerellavaginalis, Peptostreptococcus spp., Prevotella spp., Lactobacillus spp.,or Mobiluncus spp. Isolates from all bacterial strains of interest wereidentified, resulting in 39 strains in total. Isolates were frozen andstored for later use.

Example 6 Antimicrobial Susceptibility Testing

Reference type strains (8 strains total) were thawed and tested againstthe formulations described herein. After initial culture of the selectedfrozen preserved bacteria from Example 5, and 2 additional passages(sub-cultures)−(total=3 passages) on blood agar and colony suspension insaline, the saline suspension was used to inoculate the surface ofBrucella Agar with 5% Sheep Blood. Small (6 mm) discs with 10 μL offormulations F1, F2, or F3 and control gel formulation were centered onthe agar plates and incubated at 35° C. under anaerobic conditions. Thediameter of the inhibition zone in mm was measured after 48 to 96 hoursof incubation, depending on bacterial strains. Testing was conducted intriplicate.

Testing showed that a specific Lactobacillus jensenii strain andMobiluncus curtisii subsp. Holmesii strain were not affected by thepresence of formulations F1, F2, and F3. But B. fragilis, G. vaginalis,L. crispatus, M. mulieris, P. anaerobius, and P. bivia were eachinhibited by the formulations described herein, with inhibition zonesvarying from about 20 to about 60 mm. No inhibition was observed usingcontrol formulations F1, F2, and F3 prepared without tinidazole.

Example 7 MIC Determination

Minimum inhibitory concentrations were determined for 39 clinicalisolates and 8 reference strains as described below.

A hemin stock solution (5 mg/mL) was prepared by dissolving 0.1 g ofhemin in 2 mL of 1.0 N NaOH. Next, the volume was adjusted to 20 mL withdistilled water and the mixture was sterilized at 121° C. for 15minutes.

A stock vitamin K solution (10 mg/mL) was prepared by combining 0.2 mLof vitamin K₁ with 20 ml of 95% ethanol. A working solution (1 mg/ml)was prepared by combining 1 ml of the stock vitamin K solution with 9 mlsterile deionized water.

Laked sheep blood was prepared by freezing defibrinated sheep blood at−80° C. The blood was then thawed slowly at 2 to 8° C. overnight. Thethawed blood was mixed thoroughly before use and warmed in a 48 to 50°C. water bath immediately before adding to a molten brucella agar base.

Serial dilutions of formulations F1 (1.5% tinidazole), F2 (1.25%tinidazole), and F3 (1.0% tinidazole) were prepared at the followingconcentrations—F3: 1,000 μg/ml; 500 μg/ml; 250 μg/ml; 125 μg/ml; 62.5μg/ml; 31.25 μg/ml; and 15.63 μg/ml; F2: 1,250 μg/ml; 625 μg/ml; 312.5μg/ml; 156.25 μg/ml; 78.125 μg/ml; 39.06 μg/ml; and 19.53 μg/ml; F1:1,500 μg/ml; 750 μg/ml; 375 μg/ml; 187.5 μg/ml; 93.75 μg/ml; 46.875μg/ml; 23.44 μg/ml; and 11.72 μg/ml.

Brucella laked blood agar with vitamin K₁ and hemin was prepared asfollows. 28 g of brucella broth powder was combined with 15 g of agarand 1 ml each of the hemin stock solution and vitamin K working solutionin 1 L distilled water. The solution was boiled to dissolve the agar andsubsequently autoclaved at 121° C. for 15 minutes. The resulting agarsolution was then cooled to about 50° C. and 50 ml of sterile lakedsheep blood was added. Next, 2 ml of each of a given serial dilution offormulation F1, F2, or F3 (as described above) was combined with 18 mlof the laked sheep blood containing agar solution. The resultingsolution was then added to a 15×100 mm Petri dish and allowed tosolidify and prereduce in an anaerobic chamber for at least 24 hours.This gave tinidazole infused agar plates having tinidazoleconcentrations of 100 μg/ml, 50 μg/ml, 25 μg/ml, 12.5 μg/ml, 6.25 μg/ml,3.125 μg/ml, and 1.563 μg/ml (F3); 125 μg/ml, 62.5 μg/ml, 31.25 μg/ml,15.63 μg/ml, 7.81 μg/ml, 3.91 μg/ml, and 1.95 μg/ml (F2); and 150 μg/ml,75 μg/ml, 37.5 μg/ml, 18.75 μg/ml, 9.37 μg/ml, 4.69 μg/ml, and 2.34μg/ml (F1).

As in Example 6, cultures of the frozen preserved bacteria from Example5, were grown on blood agar for 24 to 96 hours. Colonies of similarmorphology were selected and suspended in prereduced brucella broth toachieve a turbidity equivalent to 0.5 McFarland.

The resulting suspension was then used to inoculate the tinidazoleinfused and control agar plates in triplicate. MIC was determined bycomparing control plates (not infused with tinidazole) against each ofthe inoculated tinidazole-infused plates. MIC was read as the minimumconcentration at which a marked reduction in bacterial growth wasobserved. The data is reported in Table 5.

TABLE 5 Bacteria MIC (μg/ml) Bacteroides fragilis Less than about 5, andin certain instances, less than about 1.6 Gardnerella vaginalis Fromless than about 2 to about 150; about 2 to about 125; or about 2 toabout 100 Lactobacillus crispatus Less than about 50, and in certaininstances, less than about 30, or less than about 25 (reference strain)Mobiluncus mulieris From about 15 to about 25 Peptostreptococcusanaerobius, Less than about 2 Prevotella bivia, Prevotella disiens, andPrevotella timonensis

The MICs for each identified strain of Lactobacillus gasseri,Lactobacillus iners, Lactobacillus jensenii, Lactobacillus vaginalis,and Mobiluncus curtisii were each greater than 150 μg/ml. Thenon-reference strain of Lactobacillus crispatus was similarly unaffectedby tinidazole at the highest concentration tested. This datademonstrates that these strains, and in particular the lactobacillusstrains which are important for vaginal health, are not inhibited bytinidazole at concentrations sufficient to inhibit bacterial strainsassociated with bacterial vaginosis.

In addition to testing the formulations including tinidazole as notedabove, formulations equivalent to formulations F1, F2, and F3 wereprepared without tinidazole to determine the anti-microbial activityattributable to the components of the F1, F2, and F3 formulations. Thesetinidazole-free formulations were appropriately diluted and testedagainst the various identified bacterial strains. No inhibition ofgrowth for any bacterial isolate was observed using the tinidazole freeformulations.

Example 8 Franz Cell Diffusion Testing

Formulations F1, F2, and F3 were subjected to Franz cell diffusiontesting. In the Franz cell (PermeGear, Inc. Hellertown, Pa.), a 0.40 μmpore polycarbonate membrane (Fisher Scientific, Pittsburgh, Pa.) wasused. The membrane area was 1.77 cm². A 480 μL sample of each testformulation was loaded in the donor cell. The receptor cell contained 8mL of 0.05 M citrate buffer, pH 3.3 (“receiver fluid”). At 0.5, 1, 2, 3,4, and 6 hours (n=6 replicates), a 500 μl sample was removed from thereceptor cell and replaced with 500 μL of fresh buffer. The study wasconducted at 37° C. Concentration of tinidazole in the receiver fluidremoved from the Franz cell was determined by LC-MS/MS.

Drug concentration per unit membrane area was plotted against the squareroot of time in hours. FIG. 15 shows the mean±standard deviation oftinidazole released (μmol)/cm² versus square root of time in hours.

There was no gel left in any of the chambers at the end of 6 hours. Dueto tinidazole saturation in the receiver chambers occurring around 180minutes, all in vitro release rates (slopes) were calculated using onlythe linear portion of the curve (R²≧0.9, i.e., samples up to andincluding 180 minutes). Rate and recovery data are shown in Table 6,below.

TABLE 6 Rate^(a) % Recovery^(a) Formulation ([μmol/cm²/√t(h)]) (at 6hours) F3 6.04 ± 0.893 98.5 ± 5.34 F2 7.56 ± 1.32  96.5 ± 4.24 F1 8.70 ±0.964 89.6 ± 4.51 ^(a)= Mean ± Standard Deviation

As can be seen in Table 6, release rate increased as tinidazoleconcentration increased. Based on 90% confidence intervals computed forthe ratios of the median in vitro release rate for pairs of formulationsF1, F2, and F3 (e.g. F1 and F2; F1 and F3; F2 and F3), the F1 and F2comparison showed these formulations can be considered to haveequivalent release rates (default “no difference” 90% confidenceintervals limits were 75% to 133.33%). The statistical equivalence ofrelease rates of the F2 and F3 formulations was not determined by thepresent comparison, while the difference in release rate between F1 andF3 was statistically significant.

Example 9 Flux and Tissue Association Using EpiVaginal™ Tissue

EpiVaginal™ tissue (produced from normal human-derivedvaginal-ectocervical epithelial cells) full thickness tissues(VEC-100-FT) on polycarbonate membranes (NUNC® single well tissueculture plate, pore size 0.4 μm) and culture medium (VEC-100-ASY) werepurchased from MatTek Corporation (Ashland, Mass.).

The culture medium (VEC-100-ASY) was pre-warmed at 37° C. Under sterileconditions and using sterile forceps, the EpiVaginal™ tissues weretransferred into 12-well plates containing the pre-warmed culturemedium. Then, the 12-well plates containing the EpiVaginal™ tissuesamples were placed in a humidified incubator (37±2° C., 5±1% CO₂) forovernight recovery. The next morning, the culture medium in the plateswas aspirated, and the EpiVaginal™ tissues were transferred into new24-well plates.

An aliquot (100 μL) of one of formulation F1, F2, or F3 was added to theapical (AP) side of EpiVaginal™ tissue in the culture insert (see FIG.16), and 500 μL, of the culture medium was added to the basolateral (BL)side of the insert. For a control group, an aliquot (100 μL) of culturemedium was added to the AP side of the insert, and 500 μL, of culturemedium was added to the BL side (the “Mediuim” in FIG. 16). All dosingwas performed under sterile conditions in a laminar flow hood.

The plates containing the tissue were incubated in a humidifiedincubator (37±1° C., 5±1% CO₂) during the study and removed only forassays at 1, 2, 4, 6, 8, 24, or 48 hours. Four replicates were studiedat each time point, for a total of 28 samples per formulation. The notedstudies were run in parallel such that plates assayed at a given timepoint were not assayed at any other time point.

At the completion of each incubation time point, the 500 μL of culturemedium on the BL side was analyzed for tinidazole content usingLC-MS/MS. The tinidazole concentrations recorded at the 1, 2, and 4 hourtime points were used to calculate the mean flux rate in EpiVaginal™tissue culture.

Simultaneously, the EpiVaginal™ tissue culture was removed from thereactor and the contents in both the AP and BL sides were carefullyaspirated and the tissues were washed twice with assay media (300 μL) toremove residual formulations. The rinses were discarded and the tissuewas gently blotted on a Kim wipe and weighed. The tissues were stored at−80° C. until further analysis, which comprised homogenization with theresulting tissue lysates analyzed by LC-MS/MS to determine the tissuetinidazole concentration at each time point.

TABLE 7 Mean Peak Tinidazole Concentration in EpiVaginal ™ Mean^(a)Tinidazole Mean Flux Tissue at Receiver Fluid Concentration Rate 1 Hour(nM) Formulation (nmol/cm²/min) (ng/mg)^(b) 1-Hour 2-Hour 4-Hour F3   18± 0.993 2,017 ± 161 1,144,500 ± 149,089 2,290,000 ± 303,425 4,400,000 ±199,499 F2 22.5 ± 2.31 2,762 ± 533 1,517,500 ± 207,746 2,855,000 ±245,289 5,567,500 ± 279,687 F1 19.0 ± 3.52 3,434 ± 498 1,735,000 ±270,000 2,627,500 ± 257,472 5,105,000 ± 713,559 ^(a)n = 4 replicates pertreatment at 1-, 2-, and 4-hour time point. ^(b)Mean peak EpiVaginal ™tissue concentrations occurred at the 1-hour time point for alltreatments (4 replicates).

As can be seen in Table 7, the mean tinidazole tissue flux ratesdiffered by less than 20% between formulations F1, F2, and F3, implyingthat flux rates were not concentration dependent. Even though flux rateswere not concentration dependent, concentration-dependent increases inthe mean maximal EpiVaginal™ tissue-associated tinidazole concentrationswere observed at the 1 hour incubation time point. All of the maximaltissue-associated tinidazole concentrations exceeded 2,000 ng/mg oftissue across the 3 formulations evaluated. Additionally, for each offormulations F1, F2, and F3, the receiver fluid mean tinidazoleconcentrations increased approximately proportionally with incubationtimes of up to 4 hours duration.

It is to be understood that the phraseology or terminology herein is forthe purpose of description and not of limitation, such that theterminology or phraseology of the present specification is to beinterpreted by the skilled artisan in light of the teachings andguidance.

The breadth and scope of the present invention should not be limited byany of the above-described exemplary embodiments, but should be definedonly in accordance with the following claims and their equivalents.

What is claimed is:
 1. A formulation suitable for the intravaginaldelivery of tinidazole to a subject in need thereof, the formulationcomprising water, tinidazole, a thermoreversible gelling agent, one ormore pharmaceutically acceptable C₁-C₇ alcohols, a solubility enhancer,and, optionally, one or more preservatives.
 2. The formulation of claim1, wherein the thermoreversible gelling agent is triblock copolymerhaving a central hydrophobic block flanked on each side with ahydrophilic block.
 3. The formulation of claim 2, wherein thehydrophobic block is polypropylene oxide).
 4. The formulation of claim3, wherein the hydrophilic block is poly(ethylene oxide).
 5. Theformulation of any one of claims 1-4, wherein the thermoreversiblegelling agent is poloxamer
 407. 6. The formulation of any one of claims1-5, wherein the one or more pharmaceutically acceptable C₁-C₇ alcoholscomprise a first pharmaceutically acceptable C₁-C₇ alcohol and a secondpharmaceutically acceptable C₁-C₇ alcohol.
 7. The formulation of claim6, wherein the first and second pharmaceutically acceptable C₁-C₇alcohols are independently selected from the group consisting ofmethanol, ethanol, isopropanol, propylene glycol,2-(2-ethoxyethoxy)ethanol, benzyl alcohol, and combinations thereof. 8.The formulation of any one of claim 6 or 7, wherein the firstpharmaceutically acceptable C₁-C₇ alcohol is benzyl alcohol.
 9. Theformulation of claims 6-8, wherein the second pharmaceuticallyacceptable C₁-C₇ alcohol is isopropanol.
 10. The formulation of claim 7or 9, wherein the isopropanol is a 60% (w/w) solution in water.
 11. Theformulation of any one of claims 1-10, wherein the solubility enhanceris selected from the group consisting of mono- and di-alkyl ethers ofisosorbide.
 12. The formulation of claim 11, wherein the mono- ordi-alkyl ether of isosorbide is dimethyl isosorbide.
 13. The formulationof any one of claims 1-10, the water comprises a mixture of deionizedwater and a buffered aqueous solution.
 14. The formulation of claim 13,wherein the buffered aqueous solution is citrate buffer.
 15. Theformulation of any one of claims 1-14, wherein the formulation has abuffered pH selected from the group consisting of about 3 to about 5,about 3.5 to about 4.5, and about 4 to about 4.5.
 16. The formulation ofany one of claims 1-15, wherein the tinidazole comprises from about 0.1to about 2% (w/w) of the formulation.
 17. The formulation of any one ofclaims 1-16, wherein the tinidazole comprises about 1%, about 1.25%, orabout 1.5% (w/w) of the formulation.
 18. The formulation of claim 1,wherein the formulation comprises about 20% poloxamer 407 (w/w), about49.6% citrate buffer (w/w), about 3.1% benzyl alcohol (w/w), about 12%(w/w) of a 60% (w/w) solution of isopropanol in water, about 13.8%dimethylisosorbide (w/w), and about 1.5% tinidazole (w/w).
 19. Theformulation of claim 1, wherein the formulation comprises about 19%poloxamer 407 (w/w), about 53% citrate buffer (w/w), about 2.9% benzylalcohol (w/w), about 11% (w/w) of a 60% (w/w) solution of isopropanol inwater, about 12.7% dimethylisosorbide (w/w), and about 1.25% tinidazole(w/w).
 20. The formulation of claim 1, wherein the formulation comprisesabout 18% poloxamer 407 (w/w), about 57% citrate buffer (w/w), about2.5% benzyl alcohol (w/w), about 10% of a 60% (w/w) solution ofisopropanol in water, about 11.5% dimethylisosorbide (w/w), and about 1%tinidazole (w/w).
 21. A method for treating bacterial vaginosis,trichomoniasis, or other disease of the vaginal cavity susceptible totinidazole, comprising intravaginally administering to a subject in needthereof a formulation comprising water, tinidazole, a thermoreversiblegelling agent, one or more pharmaceutically acceptable C₁-C₇ alcohols, asolubility enhancer, and, optionally, one or more preservatives.
 22. Themethod of claim 21, wherein the thermoreversible gelling agent istriblock copolymer having a central hydrophobic block flanked on eachside with a hydrophilic block.
 23. The method of claim 22, wherein thehydrophobic block is poly(propylene oxide).
 24. The method of claim 23,wherein the hydrophilic block is poly(ethylene oxide).
 25. The method ofclaim 22, wherein the thermoreversible gelling agent is poloxamer 407.26. The method of any one of claims 21-25, wherein the one or morepharmaceutically acceptable C₁-C₇ alcohols comprises a firstpharmaceutically acceptable C₁-C₇ alcohol and a second pharmaceuticallyacceptable C₁-C₇ alcohol.
 27. The method of claim 26, wherein the firstand second pharmaceutically acceptable C₁-C₇ alcohols are selected fromthe group consisting of methanol, ethanol, isopropanol, propyleneglycol, 2-(2-ethoxyethoxy)ethanol, benzyl alcohol, and combinationsthereof.
 28. The method of claim 26 or 27, wherein the firstpharmaceutically acceptable C₁-C₇ alcohol is benzyl alcohol.
 29. Themethod of any one of claims 26-28, wherein the second pharmaceuticallyacceptable C₁-C₇ alcohol is isopropanol.
 30. The method of claim 27 or29, wherein the isopropanol is a 60% (w/w) solution in water.
 31. Themethod of any one of claims 21-30, wherein the solubility enhancer isselected from the group consisting of mono- and di-alkyl ethers ofisosorbide.
 32. The method of claim 31, wherein the mono- or di-alkylether of isosorbide is dimethyl isosorbide.
 33. The method of any one ofclaims 21-32, wherein the water comprises a mixture of sterile deionizedwater and a buffered aqueous solution.
 34. The method of claim 33,wherein the buffered aqueous solution is citrate buffer.
 35. The methodof any one of claims 21-34, wherein the formulation has a buffered pHselected from the group consisting of about 3 to about 5, about 3.5 toabout 4.5, and about
 4. 36. The method of any one of claims 21-35wherein the tinidazole comprises from about 0.1% to about 2% (w/w) ofthe formulation.
 37. The method of any one of claims 21-36, wherein thetinidazole comprises about 1%, about 1.25%, or about 1.5% (w/w) of theformulation.
 38. The method of any one of claims 21-27, wherein thesubject is a human.
 39. A pharmaceutical composition comprising: about18-20% poloxamer 407 (w/w), about 49.6-57% citrate buffer (w/w), about2.5-3.1% benzyl alcohol (w/w), about 10-12% (w/w) of a 60% (w/w)solution of isopropanol in water, about 11.5%-13.8% dimethylisosorbide(w/w), and about 1-1.5% tinidazole (w/w) or an active metabolitethereof, for treating bacterial vaginosis, trichomoniasis, or otherdisease of the vaginal cavity susceptible to tinidazole.
 40. Apharmaceutical composition consisting of: about 18-20% poloxamer 407(w/w), about 49.6-57% citrate buffer (w/w), about 2.5-3.1% benzylalcohol (w/w), about 10-12% (w/w) of a 60% (w/w) solution of isopropanolin water, about 11.5%-13.8% dimethylisosorbide (w/w), and about 1-1.5%tinidazole (w/w) or an active metabolite thereof, for treating bacterialvaginosis, trichomoniasis, or other disease(s) of the vaginal cavitysusceptible to tinidazole.
 41. The pharmaceutical composition of claim39 or 40, wherein said pharmaceutical composition exists as a viscousliquid at room temperature and gels at or close to body temperature. 42.Use of the formulation of any one of claims 1-20 for treating bacterialvaginosis, trichomoniasis, or other disease(s) of the vaginal cavitysusceptible to tinidazole.
 43. Use of the formulation of any one ofclaims 1-20 for the manufacture of a medicament for the treatment ofbacterial vaginosis, trichomoniasis, or other disease of the vaginalcavity susceptible to tinidazole.
 44. A method for treating a vaginaldisorder comprising intravaginally administering to the subject aformulation containing about 0.1-2% tinidazole, or an active metabolitethereof.
 45. A method for administering tinidazole to a human subject,comprising: a. obtaining a formulation containing about 0.1-2%tinidazole, or an active metabolite thereof, wherein said formulationexists as a viscous liquid at room temperature; and b. intravaginallyadministering the formulation to a human subject, wherein theformulation gels at or close to body temperature.
 46. A formulationsuitable for the intravaginal delivery of tinidazole to a human subjectin need thereof, the formulation comprising tinidazole, wherein theformulation has a Franz cell flux of from about 2 to about 10(μmol/cm²)/√t(h).
 47. The formulation of claim 46, wherein the Franzcell flux is from about 6 to about 9 (μmol/cm²)/√t(h).
 48. A formulationsuitable for the intravaginal delivery of tinidazole to a human subjectin need thereof, the formulation comprising tinidazole, wherein theformulation has a mean flux rate in EpiVaginal™ tissue of from about 15nmol/cm²/min to about 30 nmol/cm²/min.
 49. The formulation of claim 48,wherein the mean flux rate in EpiVaginal™ tissue is from about 18nmol/cm²/min to about 22.5 nmol/cm²/min.
 50. A formulation suitable forthe intravaginal delivery of tinidazole to a human subject in needthereof, the formulations comprising tinidazole, wherein the formulationhas a mean tinidazole receiver fluid concentration at 1 hour of fromabout 0.5 mM to about 2.5 mM.
 51. The formulation of claim 50, whereinthe formulation has a mean tinidazole receiver fluid concentration at 1hour of from about 1.14 mM to about 1.74 mM.
 52. A tinidazole gelformulation suitable for the intravaginal delivery of tinidazole to asubject in need thereof, the formulation comprising water, tinidazole, athermoreversible gelling agent, one or more pharmaceutically acceptableC₁-C₇ alcohols, a solubility enhancer, and, optionally, one or morepreservatives, the tinidazole gel formulation prepared by the processof: a. dissolving the thermoreversible gelling agent in the water toform a first solution; b. combining the one or more pharmaceuticallyacceptable C₁-C₇ alcohols, the solubility enhancer, and the tinidazoleto form a second solution; and c. adding the second solution to thefirst solution.
 53. The tinidazole gel formulation of claim 52, whereinthe thermoreversible gelling agent is triblock copolymer having acentral hydrophobic block flanked on each side with a hydrophilic block.54. The tinidazole gel formulation of claim 53, wherein the hydrophobicblock is polypropylene oxide).
 55. The tinidazole gel formulation ofclaim 53, wherein the hydrophilic block is poly(ethylene oxide).
 56. Thetinidazole gel formulation of any one of claims 52-55, wherein thethermoreversible gelling agent is poloxamer
 407. 57. The tinidazole gelformulation of any one of claims 52-55, wherein the one or morepharmaceutically acceptable C₁-C₇ alcohols comprise a firstpharmaceutically acceptable C₁-C₇ alcohol and a second pharmaceuticallyacceptable C₁-C₇ alcohol.
 58. The formulation of claim 57, wherein thefirst and second pharmaceutically acceptable C₁-C₇ alcohols areindependently selected from the group consisting of methanol, ethanol,isopropanol, propylene glycol, 2-(2-ethoxyethoxy)ethanol, benzylalcohol, and combinations thereof.
 59. The tinidazole gel formulation ofany one of claim 57 or 58, wherein the first pharmaceutically acceptableC₁-C₇ alcohol is benzyl alcohol.
 60. The tinidazole gel formulation ofclaims 57-59, wherein the second pharmaceutically acceptable C₁-C₇alcohol is isopropanol.
 61. The tinidazole gel formulation of claim 58or 60, wherein the isopropanol is a 60% (w/w) solution in water.
 62. Thetinidazole gel formulation of any one of claims 52-61, wherein thesolubility enhancer is selected from the group consisting of mono- anddi-alkyl ethers of isosorbide.
 63. The tinidazole gel formulation ofclaim 62, wherein the mono- or di-alkyl ether of isosorbide is dimethylisosorbide.
 64. The tinidazole gel formulation of any one of claims52-63, wherein the water comprises a mixture of deionized water and abuffered aqueous solution.
 65. The tinidazole gel formulation of claim64, wherein the buffered aqueous solution is citrate buffer.
 66. Thetinidazole gel formulation of any one of claims 46-65, wherein theformulation has a pH of from about 3 to about
 5. 67. The tinidazole gelformulation of claim 66, wherein the formulation has a pH of from about3.5 to about 4.5.
 68. A process for the preparation of a tinidazole gelformulation suitable for the intravaginal delivery of tinidazole to ahuman subject in need thereof, the process comprising: a. dissolving athermoreversible gelling agent in water to form a first solution; b.combining one or more pharmaceutically acceptable C₁-C₇ alcohols, asolubility enhancer, and tinidazole to form a second solution; and c.adding the second solution to the first solution.
 69. The process ofclaim 68, wherein the thermoreversible gelling agent is triblockcopolymer having a central hydrophobic block flanked on each side with ahydrophilic block.
 70. The process of claim 69, wherein the hydrophobicblock is polypropylene oxide).
 71. The process of claim 69, wherein thehydrophilic block is poly(ethylene oxide).
 72. The process of any ofclaims 68-71, wherein the thermoreversible gelling agent is poloxamer407.
 73. The process of any of claims 68-72, wherein the one or morepharmaceutically acceptable C₁-C₇ alcohols comprise a firstpharmaceutically acceptable C₁-C₇ alcohol and a second pharmaceuticallyacceptable C₁-C₇ alcohol.
 74. The process of claim 73, wherein the firstand second pharmaceutically acceptable C₁-C₇ alcohols are independentlyselected from the group consisting of methanol, ethanol, isopropanol,propylene glycol, 2-(2-ethoxyethoxy)ethanol, benzyl alcohol, andcombinations thereof.
 75. The process of claim 73 or 74, wherein thefirst pharmaceutically acceptable C₁-C₇ alcohol is benzyl alcohol. 76.The process of claims 73-75, wherein the second pharmaceuticallyacceptable C₁-C₇ alcohol is isopropanol.
 77. The process of claim 74 or76, wherein the isopropanol is a 60% (w/w) solution in water.
 78. Theprocess of any of claims 64-77, wherein the solubility enhancer isselected from the group consisting of mono- and di-alkyl ethers ofisosorbide.
 79. The process of claim 78, wherein the mono- or di-alkylether of isosorbide is dimethyl isosorbide.
 80. The process of any ofclaims 64-79, wherein the water comprises a mixture of deionized waterand a buffered aqueous solution.
 81. The process of claim 80, whereinthe buffered aqueous solution is citrate buffer.
 82. The formulation ofclaim 50, wherein the formulation has a mean tinidazole receiver fluidconcentration at 2 hours of from about 1.5 mM to about 3.5 mM.
 83. Theformulation of claim 82, wherein the formulation has a mean tinidazolereceiver fluid concentration at 4 hours of from about 3 mM to about 6mM.
 84. The formulation of claim 82, wherein the formulation has a meantinidazole receiver fluid concentration at 2 hours of from about 2.3 mMto about 2.6 mM.
 85. The formulation of claim 83, wherein theformulation has a mean tinidazole receiver fluid concentration at 4hours of from about 4 mM to about 5.5 mM.
 86. A tinidazole gelformulation prepared by the process of any one of claims 68-81.
 87. Thetinidazole gel formulation of claims 1-20, wherein the formulation doesnot inhibit or only minimally inhibits lactobacillus strains at atinidazole concentration sufficient to inhibit Gardnerella vaginalisstrains associated with bacterial vaginosis.